Subtelomeric 5-enolpyruvylshikimate-3-phosphate synthase copy number variation confers glyphosate resistance in Eleusine indica

Genomic structural variation (SV) has profound effects on organismal evolution; often serving as a source of novel genetic variation. Gene copy number variation (CNV), one type of SV, has repeatedly been associated with adaptive evolution in eukaryotes, especially with environmental stress. Resistance to the widely used herbicide, glyphosate, has evolved through target-site CNV in many weedy plant species, including the economically important grass, Eleusine indica (goosegrass); however, the origin and mechanism of these CNVs remain elusive in many weed species due to limited genetic and genomic resources. To study this CNV in goosegrass, we present high-quality reference genomes for glyphosate-susceptible and -resistant goosegrass lines and fine-assembles of the duplication of glyphosate’s target site gene 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS). We reveal a unique rearrangement of EPSPS involving chromosome subtelomeres. This discovery adds to the limited knowledge of the importance of subtelomeres as genetic variation generators and provides another unique example for herbicide resistance evolution.

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Please select the one below that is the best fit for your research. If you are not sure, read the appropriate sections before making your selection. The assembled genomes, associated GFF annotation files, and all functional annotation information are publicly available through the International Weed Genomics Consortium online database, 'Weedpedia' (https://weedpedia.weedgenomics.org/). Raw sequencing data has been submitted to the National Center for Biotechnology Information (NCBI) Sequence Read Archive (SRA); Genome resequencing data with the accession numbers SRR23364316-SRR23364331 and RNA-seq data with the accession numbers SRR23372273-SRR23372288. None.
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resistant and susceptbale individuals from two populations
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This is a binary comparison between two distinct populations. Plants were grown in the same growth space. None.
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One glyphosate-susceptible (GS) and one glyphosate-resistant (GR) population of of E. E. indica that were previously characterized by by Zhang et et al., 2021(DOI: 10.1002 were collected from Guangdong Province, China. These populations were purified and selfpollinated for increased homozygosity and consistency of of GS GS or or GR GR phenotypes. The GR GR population was confirmed to to have EPSPS copy number variation by by DNA quantitative PCR before purification. Seeds of of purified GS GS and GR GR biotypes were sown on on wet filter paper in in Petri dishes in in a climate chamber at at 28-30°C, with 12h/12h light/dark period and 70% relative humidity. The two-leaf stage seedlings were transplanted into 28 28 × 54 54 cm cm trays (50 plants per tray) filled with potting soil and grown in in a glasshouse. At At the tillering stage, about ten individuals were randomly selected each from the GS GS and GR GR E. E. indica population and characterized. Three tillers of of each plant were separated and repotted (one tiller per pot, 60 60 pots in in total). One tiller of of each plant was used for glyphosate resistance and susceptibility phenotyping, one for EPSPS CNV estimation and one for subsequent sequencing. For glyphosate resistance and susceptibility phenotyping, one regrowth tiller (three days after tiller cloning) was treated with commercial glyphosate (41% glyphosate isopropylamine salt, 400 g ai ai ha-1 for GS GS and 1600 g ai ai ha-1 for GR), and GR GR (i.e., survivors) and GS GS (i.e., killed) phenotypes were determined three weeks after treatment. EPSPS CNV was again assessed in in the GR GR population to to ensure the CNV event was still present before genomic work began. Leaf material from untreated tillers of of corresponding resistant and susceptible plants was used for genomic DNA isolation using the Plant Genomic DNA kit (Trans Gen Biotech Beijing Co., LTD). Quantitative PCR was performed using published primer pairs and methods where EPSPS copy number was compared to to the single copy acetolactate synthase (ALS) gene as as the internal reference. The untreated tiller of of a confirmed GS GS plant was used for genomic sequencing performed at at Shanghai OE OE Biotech Co., Ltd (Shanghai, China).